Selective estrogen receptor degrader

ABSTRACT

A selective estrogen receptor degrader (SERD), a compound 3-(3,5-difluorophenyl)-2-[4-[(E)-3-[3-(fluoromethyl)azetidin-1-yl]prop-1-enyl]phenyl]-4-methyl-2H-chromen-7-ol, and its S enantiomer, (2S)-3-(3,5-difluorophenyl)-2-[4-[(E)-3-[3-(fluoromethyl)azetidin-1-yl]prop-1-enyl]phenyl]-4-methyl-2H-chromen-7-ol, or pharmaceutically acceptable salts thereof. Also provided are processes for their preparation. Also provided for is the use of these compounds for the treatment of diseases which are related to modulation of estrogen receptors, such as ER-positive breast cancer.

RELATED APPLICATIONS

This application is a continuation of U.S. patent application Ser. No.16/936,022 filed on Jul. 22, 2020, which claims priority to IndianProvisional Patent Application No. IN 201921029554 filed on Jul. 22,2019, the contents of which are hereby incorporated by reference intheir entirety.

FIELD OF THE INVENTION

The present invention relates to a selective estrogen receptor degrader(SERD), a compound3-(3,5-difluorophenyl)-2-[4-[(E)-3-[3-(fluoromethyl)azetidin-1-yl]prop-1-enyl]phenyl]-4-methyl-2H-chromen-7-ol,and its S enantiomer,(2S)-3-(3,5-difluorophenyl)-2-[4-[(E)-3-[3-(fluoromethyl)azetidin-1-yl]prop-1-enyl]phenyl]-4-methyl-2H-chromen-7-ol,or pharmaceutically acceptable salts thereof. The present inventionfurther provides processes for their preparation. The present disclosurealso relates to the use of these compounds and related methods for thetreatment of diseases which are related to the modulation of an estrogenreceptor (ER), such as ER-positive breast cancer.

BACKGROUND OF THE INVENTION

Endogenous estrogen, 17β-estradiol (E2), shows a wide variety ofbiological activities in reproductive systems, bone development andturnover, and cardiovascular systems, as well as the central nervoussystem through interactions with Estrogen Receptors (ERs). ERs have beenfound to have two isoforms, ER-α and ER-β. The link between estrogen andbreast cancer growth and development has been well established.

A number of strategies to inhibit the action of endogenous estrogen inestrogen receptor positive breast cancer are in practice. These include:selective ER modulators (SERMs) such as tamoxifen, which act asselective tissue-specific antagonist of ERs in the breast; selective ERdegraders (SERD) such as fulvestrant, which promote ER turnover; andaromatase inhibitors (AI) such as exemestane (steroidal), anastrozoleand letrozole (nonsteroidal) which inhibit estrogen biosynthesis and areprimarily used for postmenopausal women with ER-positive breast cancer.Unfortunately, many women with breast cancer initially respond well totamoxifen or AI therapy but develop resistance over a period of timeduring treatment. In the resistant form of breast cancer there isevidence that pro-growth signaling pathways downstream of estrogenreceptors still play a significant role. Recently, there has beenincreasing clinical evidence that following treatment with AIs,resistance develop due to mutations in the ligand-binding domain ofER-α, rendering it constitutively active even in the absence of ligand,thus leading to resistance.

Fanning reported that the most prevalent ER-α point mutations were Y537Sand D538G, while several others were identified at significantly reducedfrequencies. Importantly, breast cancer cell-based studies revealed thatthe Y537S and D538G mutations conferred hormone-independent activationof ER-α and reduced the inhibitory potency and efficacy of clinicallyprescribed SERMs and SERDs (Fanning et al. eLife 2016; 5:e12792).

Currently, fulvestrant is considered as a first-in-class SERD.Unfortunately, significant pharmaceutical liabilities of fulvestrant,namely its requirement of an intramuscular injection of a large volume,its poor solubility, and its lack of oral bioavailability limit itswidespread use. Therefore, the development of an orally bio-availableER-antagonist, especially one with ER degrading properties, would bebeneficial to patients who have developed resistance to currentlyavailable therapies targeting ER activity. Several novel SERDs have beenrecently developed which are currently in different phases of clinicaltrials, for example, SAR-439859 (Phase II), LSZ-102 (Phase I), AZD-9496(Phase II), GDC-810 (currently discontinued), GDC-927 (currentlydiscontinued), and others. Many non-steroidal ER antagonists arereported in prior art. For instance, the U.S. Pat. Nos. 5,395,842 and6,060,503 disclose anti-estrogenic compounds and compositions.

U.S. Pat. Nos. 5,389,646, 5,407,947; European Patent No. EP470310B1 andWIPO Publication No. WO9902512A1 disclose benzopyran compounds usefulfor treatment or prevention of conditions modulated through the estrogenreceptor.

United States Patent Application Publication No. US2004034017 (assignedto Schering Aktiengesellschaft) discloses 4-fluoroalkyl-2H-benzopyranderivatives as anti-estrogens. WIPO Publication Nos. WO2011156518A2 andWO2013090829A1 (both assigned to Aragon Pharmaceuticals Inc.) disclose alarge genus of 2H-chromene derivatives as estrogen receptor modulators.

WO2013090836A1 (Assigned to Aragon pharmaceuticals, Inc.) discloses2H-chromene derivatives having a fluorinated azetidine or pyrrolidinering in the side chain as estrogen receptor modulators/ER degraders.

WO2014205136A1 and WO2014205138A1 (both assigned to Seragonpharmaceuticals, Inc.) disclose 4-methyl-2H-chromene derivatives andstereoisomers thereof having a fluoromethylazetidine group in the sidechain as estrogen receptor modulators/ER degraders.

WO2016097073A1 (assigned to F. Hoffmann-La Roche AG/Genentech, Inc.)discloses 2H-chromene derivatives having a fluoromethylazetidine groupor a fluoromethylpyrrolidine group in the side chain as estrogenreceptor modulators/ER degraders.

WO2016189011A1 (assigned to F. Hoffmann-La Roche AG/Genentech, Inc.)discloses 2H-chromene derivatives having a fluoromethylazetidine or apyrrolidine group in the side chain as estrogen receptor modulators/ERdegraders.

WO2013090921A1 and WO2014203132A1 (both assigned to OlemaPharmaceuticals, Inc.) disclose benzopyran derivatives havingmethylpyrrolidine in the side chain as antiestrogens.

WO2016174551A1 (assigned to Pfizer Inc.) discloses 2H-chromenederivatives having N-alkylated azetidine in the side chain asanti-estrogens.

SUMMARY OF THE INVENTION

The present invention provides a compound3-(3,5-difluorophenyl)-2-[4-[(E)-3-[3-(fluoromethyl)azetidin-1-yl]prop-1-enyl]phenyl]-4-methyl-2H-chromen-7-olrepresented by Formula I:

or a pharmaceutically acceptable salt thereof.

The present invention also provides a compound(2S)-3-(3,5-difluorophenyl)-2-[4-[(E)-3-[3-(fluoromethyl)azetidin-1-yl]prop-1-enyl]phenyl]-4-methyl-2H-chromen-7-olrepresented by Formula Ia:

or a pharmaceutically acceptable salt thereof.

The compounds of present invention are selective estrogen receptordegraders and can be used for the treatment of diseases which arerelated to the modulation of ER.

Glossary

“Pharmaceutically acceptable salt” as used herein includes one or moretypes of acid addition salts formed with either organic or inorganicacids. Suitable pharmaceutically acceptable salts of the compoundsdisclosed herein include, but are not limited to, acid addition saltswhich may be salts of inorganic acids such as hydrochloric acid,hydrobromic acid, and phosphoric acid, or of organic acids such as, forexample, acetic acid, benzenesulfonic acid, methanesulfonic acid,benzoic acid, citric acid, glycolic acid, lactic acid, fumaric acid,succinic acid, adipic acid, pimelic acid, suberic acid, azelaic acid,malic acid, tartaric acid, and amino acids such as glutamic acid oraspartic acid.

The term “effective amount” as used herein refers to an amount of thecompound which is sufficient, upon single or multiple doseadministration(s) to a subject, in curing, alleviating, relieving orpartially addressing the clinical manifestation of a given disease orstate and its complications beyond that expected in the absence of suchtreatment. Thus, the result can be reduction and/or alleviation of thesigns, symptoms, or causes of a disease, or any other desired alterationof a biological system. It is understood that “a therapeuticallyeffective amount” can vary from subject to subject depending on age,weight, general condition of the subject, the condition being treated,the severity of the condition being treated, and the judgment of theprescribing physician.

The term “treating” or “treatment” as used herein refer to completely orpartially curing, alleviating, ameliorating, improving, relieving,delaying onset of, inhibiting progression of, reducing severity of,and/or reducing incidence of one or more symptoms or features of aparticular disease, disorder, and/or condition.

A human in need of the methods or compounds or treatments disclosedherein are those who are either suffering from the particular disease,disorder, and/or conditions described herein or at a recognized risk,such as by medical diagnosis, of developing that particular disease,disorder, and/or condition.

BRIEF DESCRIPTION OF DRAWINGS

FIG. 1. In-vivo efficacy of a compound of Formula I and a compound ofFormula Ia in MCF7-Y537S mice xenografts.

DETAILED DESCRIPTION OF THE INVENTION

Previously the present applicant had filed a patent applicationpublished as WIPO Publication No. WO2017072792A1 covering 2H-chromenederivatives having a heterocyclic ring in the side chain as ERantagonists/degraders. The present inventors, in a quest to furtherdevelop a better orally bio-available ER-antagonist especially with ERdegrading properties, surprisingly found that the compound of Formula Ihaving a 7-hydroxy chromene moiety and an azetidine ring side chain inthe structure exhibits a significant degradation of estrogen receptors.It was further found that the S-stereoisomer of the compound of FormulaI was significantly more potent than the R-isomer. Moreover, it wassurprisingly found that the S-isomer of the compound of Formula Iexhibits pharmacokinetic properties which make it much more efficaciousover the R-isomer. Accordingly, in one aspect, the present inventionprovides a compound3-(3,5-difluorophenyl)-2-[4-[(E)-3-[3-(fluoromethyl)azetidin-1-yl]prop-1-enyl]phenyl]-4-methyl-2H-chromen-7-olrepresented by Formula I:

or a pharmaceutically acceptable salt thereof.

When the present inventors carried out the chiral resolution of thecompound of Formula I into its enantiomers, the ‘S’ enantiomer wassurprisingly found to be significantly superior to the ‘R’ enantiomer interms of both its in-vitro potency in an MCF-7 growth inhibitionassay/ER-α degradation assay as well as in its pharmacokinetic profile.Accordingly in a second aspect, the present invention provides acompound(2S)-3-(3,5-difluorophenyl)-2-[4-[(E)-3-[3-(fluoromethyl)azetidin-1-yl]prop-1-enyl]phenyl]-4-methyl-2H-chromen-7-olrepresented by Formula Ia:

and/or a pharmaceutically acceptable salt thereof.

One aspect of the present invention is therefore a composition in whichat least 75% of the total amount of enantiomers of Formula I that arepresent in the composition are the S enantiomer.

In certain embodiments, this percentage may be at least 85%, at least90%, at least 95%, at least 99% and 100% of the enantiomers of Formula Ithat are present are the S enantiomer. In other embodiments, thecomposition does not contain the R enantiomer of Formula I.

In another aspect, the present invention provides a compound of FormulaIa or a pharmaceutically acceptable salt thereof, which is substantiallyfree of compound of Formula Ib

The term “substantially free of compound of Formula Ib” refers tocontent of compound of Formula Ib which is less than 25%, less than 20%,less than 15%, less than 10%, less than 9%, less than 8%, less than 7%,less than 6%, less than 5%, less than 4%, less than 3%, less than 2%,less than 1%, less than 0.9%, less than 0.8%, less than 0.7%, less than0.6%, less than 0.5%, less than 0.4%, less than 0.3%, less than 0.2%,less than 0.1%, less than 0.09%, less than 0.05% or less than 0.01% w/wwith respect to compound of Formula Ia or the compound of Formula Ib isabsent.

Thus in one embodiment, the present invention provides a compound ofFormula Ia or a pharmaceutically acceptable salt thereof, wherein thecontent of compound of Formula Ib is less than 25%, less than 20%, lessthan 15%, less than 10%, less than 5%, less than 1%, less than 0.5%,less than 0.1%, less than 0.05%, less than 0.01% w/w or absent withrespect to the compound of Formula Ia.

In another embodiment, the present invention provides a compound ofFormula Ia or a pharmaceutically acceptable salt thereof, wherein theenantiomeric ratio of the compound of Formula Ia to the compound ofFormula Ib is greater than 75:25, greater than 80:20, greater than85:15, greater than 90:10, greater than 95:5, greater than 96:4, greaterthan 97:3, greater than 98:2, greater than 99:1 or is 100:0.

In another embodiment, the present invention provides a compound ofFormula Ia or a pharmaceutically acceptable salt thereof, wherein theenantiomeric ratio of the compound of Formula Ia to the compound ofFormula Ib is greater than 80:20. In another embodiment, theenantiomeric ratio of the compound of Formula Ia is greater than 85:15.In another embodiment, the enantiomeric ratio of the compound of FormulaIa is greater than 90:10. In another embodiment, the enantiomeric ratioof the compound of Formula Ia is greater than 95:5. In anotherembodiment, the enantiomeric ratio of the compound of Formula Ia isgreater than 96:4. In another embodiment, the enantiomeric ratio of thecompound of Formula Ia is greater than 97:3. In another embodiment, theenantiomeric ratio of the compound of Formula Ia is greater than 98:2.In another embodiment, the enantiomeric ratio of the compound of FormulaIa is greater than 99:1. In another embodiment, the enantiomeric ratioof the compound of Formula Ia is 100:0 i.e. the compound of Formula Ib(R enantiomer) is absent.

The present invention also includes the prodrugs or deuteratedderivatives of the compound of Formula I or Formula Ia.

The compounds of the present invention as described herein areER-antagonists especially with ER degrading properties, and thereforeare believed to be useful as medicaments, particularly for the treatmentof diseases that are ER dependent or ER mediated, such as cancer,selected from, but not limited to breast cancer, ovarian cancer, braincancer and endometrial cancer.

Given the central role of ER-α in breast cancer development andprogression, the compounds of the present invention can be useful in thetreatment of breast cancer, either alone or in combination with otheragent, including but not limited to: aromatase inhibitors (such asanastrozole, letrozole, and the like), SERMs (such as tamoxifen,raloxifene, and the like), antiestrogens (such as fulvestrant and thelike), luteinizing hormone-releasing hormone (LH-RH) agonists (such asleuprolide and the like), CDK4/6 inhibitors (such as palbociclib and thelike) or other chemotherapeutic agents including anthracylines, platins,nitrogen mustard alkylating agents, and the like.

Thus, in another aspect, the present invention provides a method oftreatment of an ER dependent or ER mediated disease or condition in ahuman being in need thereof, comprising administering thereto aneffective amount of a compound of Formula I or a pharmaceuticallyacceptable salt thereof.

In another embodiment, the present invention provides a method oftreatment of an ER dependent or ER mediated disease or condition in ahuman being in need thereof, comprising administering thereto aneffective amount of a compound of Formula Ia or a pharmaceuticallyacceptable salt thereof.

In another embodiment, the present invention provides a method oftreatment of cancer selected from breast cancer, endometrial cancer,brain cancer and ovary cancer in a human being in need thereof,comprising administering thereto an effective amount of a compound ofFormula I or a pharmaceutically acceptable salt thereof.

In another embodiment, the present invention provides a method oftreatment of cancer selected from breast cancer, endometrial cancer,brain cancer and ovary cancer in a human being in need thereof,comprising administering thereto an effective amount of a compound ofFormula Ia or a pharmaceutically acceptable salt thereof.

In another embodiment, the present invention provides a method oftreatment of breast cancer, comprising administering an effective amountof a compound of Formula I or compound of Formula Ia or apharmaceutically acceptable salt thereof.

Pharmaceutical Composition

The compounds disclosed herein may be formulated into a composition thatadditionally comprises suitable pharmaceutically acceptable carriers,including excipients and other compounds that facilitate administrationof the compound to a subject. The pharmaceutical compositions of thepresent invention may be formulated in a conventional manner using oneor more pharmaceutically acceptable excipients. The pharmaceuticallyacceptable excipients may include items such as diluents, disintegrants,binders, lubricants, glidants, polymers, coating agents, solvents,cosolvents, preservatives, wetting agents, thickening agents,antifoaming agents, sweetening agents, flavouring agents, antioxidants,colorants, solubilizers, plasticizer, dispersing agents, and the like.The compounds of the present invention may be formulated in the form ofpills, tablets, coated tablets, capsules, powder, granules, pellets,patches, implants, films, liquids, semi-solids, gels, aerosols,emulsions, elixirs, and the like. Such pharmaceutical compositions andthe processes for preparing the same are described, for example, inRemington: The Science and Practice of Pharmacy (D. B. Troy, Editor,21st Edition, Lippincott, Williams & Wilkins, 2006), the contents ofwhich are incorporated herein by reference in their entirety. In certainembodiments, the compounds and compositions described herein may beadministered orally, parenterally, intramuscularly, transdermally orintravenously.

Thus, in one embodiment, the present invention provides a pharmaceuticalcomposition comprising a compound of Formula I or a compound of FormulaIa, or a pharmaceutically acceptable salt thereof, with apharmaceutically acceptable carrier, diluent, or excipient.

Suitable doses of the compounds for use in treating the diseases asdescribed herein can be determined by those skilled in the relevant art.Therapeutic doses are generally identified through a dose ranging studyin humans based on preliminary evidence derived from animal studies.Doses must be sufficient to result in a desired therapeutic benefitwithout causing unwanted side effects. The mode of administration,dosage forms, and suitable pharmaceutical excipients can also beunderstood and adjusted by those skilled in the art.

The present invention is further illustrated in detail with reference tothe following examples.

It should be understood that the examples herein are merelyillustrative, and do not limit the scope of the present disclosure orthe claims appended hereto.

Process of Preparation

A compound of Formula I, a compound of Formula Ia, and their closelyrelated analogues were prepared as described below. All solvents andreagents were used as obtained from commercial sources unless otherwiseindicated. ¹H-NMR spectra were recorded with a Bruker® spectrometeroperating at 500 MHz in deuterated DMSO.

The compound of Formula I or the compound of Formula Ia can be convertedinto their salts by methods known in the art, including, for example,dissolving a compound of Formula I or Formula Ia in a suitable solventand treating it with an appropriate acid.

Example 1: Preparation of3-(3,5-difluorophenyl)-2-{4-[(E)-3-(3-fluoromethylazetidin-1-yl)propenyl]phenyl}-4-methyl-2H-chromen-7-ol (Compound ofFormula I)

Step-I: Preparation of 2-(3,5-difluorophenyl)-1-(2,4-dihydroxyphenyl)ethanone

Oxalyl chloride (5.98 mL, 0.070 mol) was added dropwise to a stirredsolution of 3,5-difluorophenylacetic acid (10 g, 0.058 mol) andN,N-dimethylformamide (0.5 mL) in dichloromethane (100 mL) at roomtemperature and was stirred at room temperature for 30 min. The reactionmixture was concentrated under reduced pressure at 30-35° C. and thendissolved in dichloromethane (20 mL). The resultant solution was addedto a stirred solution of resorcinol (9.58 g, 0.087 mol) and aluminiumchloride (11.60 g, 0.087 mol) in dichloromethane (80 mL) at 0-5° C. andwas stirred at room temperature for 16 hours. The reaction was slowlyquenched with 2N hydrochloric acid solution (120 mL) at 0-5° C. andstirred for 1 hr at same temperature. The solid was filtered andsuccessively washed with water and n-hexane. The resulting solid wasdried under vacuum to give2-(3,5-difluorophenyl)-1-(2,4-dihydroxyphenyl)ethanone.

Step-II: Preparation of 2-(3,5-difluorophenyl)-1-[2-hydroxy-4-(tetrahydropyran-2-yloxy)phenyl]ethanone

3,4-Dihydro-2H-pyran (45.58 mL, 0.50 mol) was added to a stirredsolution of 2-(3,5-difluorophenyl)-1-(2,4-dihydroxyphenyl)ethanone (44.0g, 0.167 mol) and pyridinium p-toluene sulfonate (6.28 g, 0.025 mol) indichloromethane (880 mL) at room temperature. The reaction mixture wasstirred at room temperature for 2 hours and quenched with an aqueoussaturated solution of sodium bicarbonate. The organic layer wasseparated and the aqueous layer was again extracted withdichloromethane. The combined organic layer was dried over anhydroussodium sulfate and concentrated under reduced pressure to give crudemass which was stirred in a mixture of n-hexane:ethyl acetate (95:5) atroom temperature and filtered to give2-(3,5-difluorophenyl)-1-[2-hydroxy-4-(tetrahydropyran-2-yloxy)-phenyl]ethanone.

Step-III: Preparation of3-(3,5-difluorophenyl)-2-(4-iodophenyl)-7-(tetrahydropyran-2-yloxy)chroman-4-one

1,8-Diazabicyclo(5.4.0)undec-7-ene (DBU, 0.055 g, 0.00036 mol) was addedto a stirred slurry of2-(3,5-difluorophenyl)-1-[2-hydroxy-4-(tetrahydropyran-2-yloxy)-phenyl]ethanone(0.5 g, 0.0014 mol), 4-iodo benzaldehyde (0.37 g, 0.0016 mol) andpiperidine (0.03 g, 0.00036 mol) in isopropyl alcohol (10 mL). Thereaction mixture was heated to 90-95° C. for 3 hours. The solvent wasremoved under reduced pressure to give3-(3,5-difluorophenyl)-2-(4-iodophenyl)-7-(tetrahydropyran-2-yloxy)chroman-4-one.

Step-IV: Preparation of3-(3,5-difluorophenyl)-2-(4-iodophenyl)-4-methyl-7-(tetrahydropyran-2-yloxy)-chroman-4-ol

Methyl magnesium chloride (3M) in tetrahydrofuran (THF, 1.6 mL) wasadded to a stirred solution of3-(3,5-difluorophenyl)-2-(4-iodophenyl)-7-(tetrahydropyran-2-yloxy)-chroman-4-one(0.8 g, 0.0014 mol) in anhydrous THF (12 mL) at 20-25° C. and stirredfor 1 hour. The reaction mixture was quenched with an aqueous saturatedammonium chloride solution at 0-5° C. followed by water. The organiclayer was separated and the aqueous layer was again extracted with ethylacetate. The combined organic layer was dried over anhydrous sodiumsulfate and solvent was removed under vacuum to give the title compound.

Step-V: Preparation of3-(3,5-difluorophenyl)-2-(4-iodophenyl)-4-methyl-2H-chromen-7-ol

A solution of concentrated sulfuric acid (0.22 mL, 0.0042 mol) inmethanol (2 mL) was added to a stirred solution of3-(3,5-difluorophenyl)-2-(4-iodophenyl)-4-methyl-7-(tetrahydropyran-2-yloxy)-chroman-4-ol(0.8 g, 0.0014 mol) in methanol (10 mL) and heated at 65-70° C. for 3hours. The reaction mixture was cooled to 0-5° C. Saturated sodiumbicarbonate solution was added to the above reaction mixture and wasextracted with ethyl acetate. The combined organic layer was dried overanhydrous sodium sulfate and solvent was removed under vacuum to give acrude mass which was purified by column chromatography (silica gel,toluene:ethyl acetate (97:3)) to give the title compound.

Step-VI: Preparation of3-(3,5-difluorophenyl)-2-(4-iodophenyl)-4-methyl-7-(tetrahydrohydropyran-2-yloxy)-2H-chromene

3,4-Dihydro-2H-pyran (10.34 mL, 0.113 mol) was added to a stirredsolution of3-(3,5-difluorophenyl)-2-(4-iodophenyl)-4-methyl-2H-chromen-7-ol (18 g,0.038 mol) and pyridinium p-toluenesulfonate (1.42 g, 0.0057 mol) indichloromethane (200 mL) at room temperature and was stirred for 16 hrs.The reaction mixture was quenched with a saturated solution of sodiumbicarbonate. The organic layer was separated and the aqueous layer wasextracted with dichloromethane. The combined organic layer was driedover anhydrous sodium sulfate and solvent was removed under vacuum togive a crude mass which was purified by column chromatography (silicagel, toluene) to give the title compound.

Step-VII: Preparation of(E)-3-{4-[3-(3,5-difluorophenyl)-4-methyl-7-(tetrahydropyran-2-yloxy)-2H-chromen-2-yl]phenyl}acrylicAcid ethyl ester

Ethyl acrylate (0.23 g, 0.0022 mol) was added to a stirred solution of3-(3,5-difluorophenyl)-2-(4-iodophenyl)-4-methyl-7-(tetrahydrohydropyran-2-yloxy)-2H-chromene(0.25 g, 0.00045 mol) and triethylamine (0.37 mL, 0.0027 mol) inN-methyl-2-pyrrolidone (2 mL) followed by addition of Pd(PPh₃)₂Cl₂(0.003 g, 0.0000044 mol) at room temperature. The resultant reactionmixture was heated at 95° C. for 1 hour. Water was added to the reactionmixture and the product was extracted with ethyl acetate. The combinedorganic layer was dried over anhydrous sodium sulfate and solvent wasremoved under vacuum to give a crude mass which was purified by columnchromatography (silica gel, n-hexane: ethyl acetate (85:15)) to give thetitle compound.

Step-VIII: Preparation of(E)-3-{4-[3-(3,5-Difluorophenyl)-4-methyl-7-(tetrahydropyran-2-yloxy)-2H-chromen-2-yl]-phenyl}prop-2-en-1-ol

Diisobutylaluminium hydride (20%) solution in toluene (0.56 mL, 0.00079mol) was added to a stirred solution of(E)-3-{4-[3-(3,5-difluorophenyl)-4-methyl-7-(tetrahydropyran-2-yloxy)-2H-chromen-2-yl]-phenyl}acrylicacid ethyl ester (0.14 g, 0.00026 mol) in toluene (5.6 mL) at −30° C.and was stirred for 45 min. at −20 to −25° C. Methanol (0.5 mL) andsodium potassium tartrate (20%) solution (5 mL) was dropwise added at−20° C. The reaction mixture was brought to room temperature and wastreated with water at room temperature. The organic layer was separatedand the aqueous layer was extracted with dichloromethane. The combinedorganic layer was dried over anhydrous sodium sulfate and solvent wasremoved under reduced pressure to give a crude mass which was purifiedby column chromatography (silica gel, n-hexane:ethyl acetate (60:40)) togive the title compound.

Step-IX:1-((E)-3-{4-[3-(3,5-difluorophenyl)-4-methyl-7-(tetrahydropyran-2-yloxy)-2H-chromen-2-yl]-phenyl}allyl)-3-fluoromethylazetidine

Iodine (1.02 g, 0.0041 mol) was added portion wise to a stirred solutionof triphenyl phosphine (1.07 g, 0.0041 mol) and imidazole (0.31 g,0.0045 mol) in dichloromethane (10 mL) at 0-5° C. The reaction mixturewas stirred at 0-5° C. for 30 min. A solution of(E)-3-{4-[3-(3,5-difluorophenyl)-4-methyl-7-(tetrahydropyran-2-yloxy)-2H-chromen-2-yl]phenyl}prop-2-en-1-ol(1.0 g, 0.0020 mol) in dichloromethane (10 mL) was added at 0-5° C. tothis reaction mixture and stirred for 20 minutes. It was slowly pouredinto a cold solution of sodium bicarbonate and extracted withdichloromethane. The combined organic layer was successively washed withan aqueous solution of sodium metabisulphite and brine solution. Theorganic layer was dried over anhydrous sodium sulfate and solvent wasremoved under vacuum to give a crude mass. A mixture of n-hexane: ethylacetate (9:1) (20 mL) was added to the above crude mass and stirred for30 min at room temperature. Finally it was filtered and the filtrate wasconcentrated under reduced pressure at 35-38° C. to give a crude mass,which was again stirred in a mixture of n-hexane: ethyl acetate (9:1)(10 mL) for 30 min. It was again filtered and the filtrate wasconcentrated under reduced pressure at 35-38° C. to give3-(3,5-difluorophenyl)-2-[4-((E)-3-iodopropenyl)phenyl]-4-methyl-7-(tetrahydropyran-2-yloxy)-2H-chromene.

Triethylamine (0.56 mL, 0.004 mol) was added to a stirred solution of3-fluoromethyl azetidine hydrochloride (0.38 g, 0.003 mol) inacetonitrile (10 mL). The reaction mixture was stirred for 30 minutes atroom temperature. A solution of3-(3,5-difluorophenyl)-2-[4-((E)-3-iodopropenyl)phenyl]-4-methyl-7-(tetrahydro-pyran-2-yloxy)-2H-chromene(1.2 g, 0.002 mol) in acetonitrile (10 mL) was added to the reactionmixture at room temperature and stirring was continued for 45 minutes.Water was added to the reaction mixture and was extracted with ethylacetate. The organic layer was separated, dried over anhydrous sodiumsulfate and solvent was removed under reduced pressure to give a crudemass which was purified by column chromatography (silica gel,dichloromethane:methanol (97:3)) to give the title compound.

Step-X: Preparation of3-(3,5-difluorophenyl)-2-{4-[(E)-3-(3-fluoromethylazetidin-1-yl)propenyl]phenyl}-4-methyl-2H-chromen-7-ol (Formula I)

A solution of sulfuric acid (0.75 mL, 0.014 mol) in methanol (70 mL) wasadded to a stirred solution of1-((E)-3-{4-[3-(3,5-difluorophenyl)-4-methyl-7-(tetrahydropyran-2-yloxy)-2H-chromen-2-yl]-phenyl}allyl)-3-fluoromethylazetidine (7.6 g, 0.014 mol) in methanol (20 mL) at 0-5° C. The reactionmixture was allowed to stir for 30 minutes at room temperature. Asaturated solution of sodium bicarbonate and water were added at 0-5° C.and was extracted with dichloromethane. The combined organic layer wasdried over anhydrous sodium sulfate and concentrated under reducedpressure to get residue which was purified by column chromatography(silica gel, dichloromethane:methanol (90:10)) to give the titlecompound.

Example 2: Preparation of(2S)-3-(3,5-difluorophenyl)-2-[4-[(E)-3-[3-(fluoromethyl)azetidin-1-yl]prop-1-enyl]phenyl]-4-methyl-2H-chromen-7-ol(Compound of Formula Ia) and(2R)-3-(3,5-difluorophenyl)-2-[4-[(E)-3-[3-(fluoromethyl)azetidin-1-yl]prop-1-enyl]phenyl]-4-methyl-2H-chromen-7-ol(Compound of Formula Ib)

The enantiomers from the racemic mixture of example 1 were separated bychiral HPLC (Column: Chiralcel® OD-H (250×30 mm, 5μ); mobilephase-n-hexane:ethanol:diethyl amine 900:100:1) wherein R enantiomer(Compound of Formula Ib) was eluted first followed by desired Senantiomer (Compound of Formula Ia). Further the specific opticalrotation (SOR) of compound of Formula Ia was determined by usingfollowing test conditions:

Concentration: 1% w/v in acetone;

Temperature: 25° C.;

Source of light: Sodium lamp (D line);

SOR of Compound of Formula Ia: [α]_(D) ²⁵=+224.40°.

The chiral purity of compound of Formula Ia was determined by HPLC asper following analytical conditions:

Column: CHIRALCEL® OD-3 (250×4.6) mm 3 μm

Mobile phase: n-hexane/ethanol/diethylamine (90/10/0.1, v/v/v)

Flow rate: 1.0 mL/min; Column Temperature: 25° C.; detector: UV230 nm;

Sample concentration: 0.5 mg/ml,

Diluent: Mobile phase.

Chiral Purity of compound of Formula Ia:=99.69:0.31 (S:R); Relativeretention time (RRT) with respect to compound of Formula Ib=About 1.1

Example 3: Preparation of3-(3,5-difluorophenyl)-2-[4-[(E)-3-[3-(fluoromethyl)azetidin-1-yl]prop-1-enyl]phenyl]-4-methyl-2H-chromen-6-ol(Compound 2)

Racemic compound 2 was prepared by following an analogous process ofExample 1 (step III-step X) wherein2-(3,5-difluorophenyl)-1-(2-hydroxy-5-tetrahydropyran-2-yloxy-phenyl)ethanonewas used instead of 2-(3,5-difluorophenyl)-1-[2-hydroxy-4-(tetrahydropyran-2-yloxy)-phenyl]ethanone in stepIII.

Example 4: Preparation of3-(3,5-difluorophenyl)-2-{4-[(E)-3-(4-fluoromethylpiperidin-1-yl)propenyl]phenyl}-4-methyl-2H-chromen-7-ol(Compound 3)

Step I:1-((E)-3-{4-[3-(3,5-difluorophenyl)-4-methyl-7-(tetrahydropyran-2-yloxy)-2H-chromen-2-yl]-phenyl}allyl)-4-fluoromethylpiperidine

A solution of methanesulfonyl chloride (0.11 mL, 1.4 mmol) indichloromethane (1 mL) was added drop-wise to a stirred solution of(E)-3-{4-[3-(3,5-difluorophenyl)-4-methyl-7-(tetrahydropyran-2-yloxy)-2H-chromen-2-yl]phenyl}prop-2-en-1-ol(0.57 g, 1.17 mmol) and triethylamine (0.24 mL, 1.75 mmol) indichloromethane (5 mL) at 0-5° C. The reaction mixture was furtherstirred at 0-5° C. for 30 minutes. Water was added to the reactionmixture and the organic layer was separated. The aqueous layer wasextracted with dichloromethane. The combined organic layer was driedover anhydrous sodium sulfate and used for the next step. This solutionwas added to a solution of triethylamine (0.65 mL, 4.7 mmol) and4-fluoromethylpiperidine hydrochloride (0.54 g, 3.5 mmol) inacetonitrile (6 mL) at 0-5° C. The reaction mixture was stirred at roomtemperature for 1.5 hours. Water was added and the mixture was extractedwith dichloromethane. The combined organic layer was dried overanhydrous sodium sulfate and concentrated under reduced pressure to geta crude mass which was purified by column chromatography (silica gel,methanol:dichloromethane (5:95)) to yield the title compound.

Step II:3-(3,5-difluorophenyl)-2-{4-[(E)-3-(4-fluoromethylpiperidin-1-yl)propenyl]phenyl}-4-methyl-2H-chromen-7-ol(Compound 3)

A solution of1-((E)-3-{4-[3-(3,5-difluorophenyl)-4-methyl-7-(tetrahydropyran-2-yloxy)-2H-chromen-2-yl]-phenyl}allyl)-4-fluoromethylpiperidine(0.7 g, 1.18 mmol) in a mixture of sulfuric acid (0.07 mL) and methanol(5 mL) was stirred at room temperature for 10 minutes. The reactionmixture was made alkaline with a saturated solution of sodiumbicarbonate and extracted with dichloromethane. The combined organiclayer was dried over anhydrous sodium sulfate and concentrated underreduced pressure to get a crude mass which was purified by columnchromatography (silica gel, methanol:dichloromethane (8:92)) to get thetitle compound.

Example 5: Preparation of3-(3,5-difluorophenyl)-2-[4-[(E)-3-[(3R)-3-(fluoromethyl)pyrrolidin-1-yl]prop-1-enyl]phenyl]-4-methyl-2H-chromen-7-ol(Compound 4)

The compound 4 was prepared following an analogous process of Example 4by using (3R)-3-(fluoromethyl)pyrrolidine hydrochloride instead of4-fluoromethylpiperidine hydrochloride in step-I.

Example 6: Preparation of3-(3,5-difluorophenyl)-4-methyl-2-[4-[(E)-3-[(3R)-3-methylpyrrolidin-1-yl]prop-1-enyl]phenyl]-2H-chromen-7-ol(Compound 5)

The compound 5 was prepared by following an analogous process of Example4 by using (3R)-3-methylpyrrolidine hydrochloride instead of4-fluoromethylpiperidine hydrochloride in step-I.

Example 7: Preparation of3-(3,5-difluorophenyl)-2-{4-[(Z)-3-(3-fluoromethylazetidin-1-yl)propenyl]phenyl}-4-methyl-2H-chromen-7-ol(Compound 6)

Step I:3-[4-[3-(3,5-difluorophenyl)-4-methyl-7-tetrahydropyran-2-yloxy-2H-chromen-2-yl]phenyl]prop-2-yn-1-ol

Bis(triphenylphosphine)palladium(II) dichloride (0.125 g, 0.18 mmol) wasadded to a stirred solution of3-(3,5-difluorophenyl)-2-(4-iodophenyl)-4-methyl-7-tetrahydropyran-2-yloxy-2H-chromene(2.0 g, 3.6 mmol), propargyl alcohol (0.60 g, 10.7 mmol) and cuprous(I)iodide (0.054 g, 0.29 mmol) in a mixture oftetrahydrofuran:triethylamine (1:1, 64 mL). Stirring was continued atroom temperature for 2 hours. The reaction mixture was concentratedunder reduced pressure to get a crude residue which was purified bycolumn chromatography (silica gel, toluene-ethyl acetate (4:1)) to yieldthe title compound.

Step II:1-[3-[4-[3-(3,5-difluorophenyl)-4-methyl-7-tetrahydropyran-2-yloxy-2H-chromen-2-yl]phenyl]prop-2-ynyl]-3-(fluoromethyl)azetidine

The step II compound was prepared by following an analogous process ofExample 4, step-I.

Step III:1-[(Z)-3-[4-[3-(3,5-difluorophenyl)-4-methyl-7-tetrahydropyran-2-yloxy-2H-chromen-2-yl]phenyl]allyl]-3-(fluoromethyl)azetidine

Lindlar catalyst (0.125 g, 25% w/w) was added to a solution of1-[3-[4-[3-(3,5-difluorophenyl)-4-methyl-7-tetrahydropyran-2-yloxy-2H-chromen-2-yl]phenyl]prop-2-ynyl]-3-(fluoromethyl)azetidine (0.50 g, 0.89 mmol) and quinoline(0.05 g, 10% w/w) in ethanol (10 mL). The reaction mixture was stirredunder hydrogen atmosphere using a hydrogen gas filled balloon at roomtemperature for 5 hours. The reaction mixture was filtered and washedwith ethanol. The filtrate was concentrated under reduced pressure toget a crude mass which was purified by column chromatography (silicagel, dichloromethane:methanol (97:3)) to yield the title compound.

Step IV:3-(3,5-difluorophenyl)-2-{4-[(Z)-3-(3-fluoromethylazetidin-1-yl)propenyl]phenyl}-4-methyl-2H-chromen-7-ol(Compound 6)

THP protection was removed using an analogous process of Example 4 (stepII) to obtain the racemic Compound 6.

Example 8: Preparation of3-(3,5-difluorophenyl)-2-[4-[3-[3-(fluoromethyl)azetidin-1-yl]prop-1-ynyl]phenyl]-4-methyl-2H-chromen-7-ol (Compound 7)

The racemic compound 7 was prepared by removing the THP protection ofthe Example 7 (step II) compound using an analogous process of Example4, step II.

The ¹H NMR data of compounds of Formula I, Formula Ia, and relatedanalogues are provided below:

Compound Chirality^(#) Structure/Chemical Name ¹H NMR data Formula IRacemic

  Formula I 3-(3,5-Difluorophenyl)-2-[4-[(E)-3-[3-(fluoromethyl)azetidin-1-yl]prop-1-enyl]phenyl]-4-methyl-2H-chromen-7-ol (DMSO-d₆, 500 MHz); 2.12 (s, 3H);3.10-3.23 (m, 1H); 3.89-4.00 (m, 4H); 4.15 (t, J = 10.52 Hz, 2H); 4.55(d, J = 4.68 Hz, 1H); 4.67 (d, J = 4.68 Hz, 1H); 6.12-6.23 (m, 3H); 6.43(dd, J₁ = 8.44 Hz, J₂ = 2.44 Hz, 1H); 6.80 (d, J = 15.97 Hz, 1H);7.06-7.20 (m, 3H); 7.25 (d, J = 8.44 Hz, 1H); 7.36 (d, J = 8.28 Hz, 2H);7.43 (d, J = 8.28 Hz, 2H); one exchangeable proton. Formula Ia S

  Formula Ia (2S)-3-(3,5-Difluorophenyl)-2-[4-[(E)-3-[3-(fluoromethyl)azetidin-1-yl]prop-1-enyl]phenyl]-4-methyl-2H-chromen-7-ol (DMSO-d₆, 500 MHz); 2.12 (s, 3H);2.73-2.85 (m, 1H); 3.05 (t, J = 6.65 Hz, 2H); 3.21 (d, J = 5.50 Hz, 2H);3.36 (t, J = 7.25 Hz, 2H); 4.51 (d, J = 6.05 Hz, 1H); 4.60 (d, J = 5.80Hz, 1H); 6.11 (s, 1H); 6.14-6.21 (m, 2H); 6.43 (dd, J₁ = 8.40 Hz, J₂ =2.40 Hz, 1H); 6.50 (d, J = 15.95 Hz, 1H); 7.04-7.09 (m, 2H); 7.11-7.17(m, 1H); 7.24 (d, J = 7.24 Hz, 1H); 7.29 (d, J = 8.25 Hz, 2H); 7.35 (d,J = 8.25 Hz, 2H); one exchangeable proton. Formula Ib R

  Formula Ib (2R)-3-(3,5-Difluorophenyl)-2-[4-[(E)-3-[3-(fluoromethyl)azetidin-1-yl]prop-1-enyl]phenyl]-4-methyl-2H-chromen-7-ol (DMSO-d₆, 500 MHz); 2.12 (s, 3H);2.73-2.84 (m, 1H); 3.03 (t, J = 6.65 Hz, 2H); 3.20 (d, J = 5.60 Hz, 2H);3.35 (t, J = 7.30 Hz, 2H); 4.51 (d, J = 6.10 Hz, 1H); 4.60 (d, J = 6.10Hz, 1H); 6.11 (s, 1H); 6.14-6.21 (m, 2H); 6.43 (dd, J₁ = 8.40 Hz, J₂ =2.35 Hz, 1H); 6.50 (d, J = 16.00 Hz, 1H); 7.04-7.10 (m, 2H); 7.11-7.17(m, 1H); 7.24 (d, J = 8.45 Hz, 1H); 7.29 (d, J = 8.25 Hz, 2H); 7.35 (d,J = 8.20 Hz, 2H); one exchangeable proton. Compound 2 Racemic

  Compound 2 3-(3,5-Difluorophenyl)-2-[4-[(E)-3-[3-(fluoromethyl)azetidin-1-yl]prop-1-enyl]phenyl]-4-methyl-2H-chromen-6-ol (DMSO-d₆, 500 MHz); 2.12 (s, 3H);2.78-2.91 (br m, 1H); 3.14-3.25 (m, 2H); 3.28-3.38 (m, 2H); 3.45-3.58(m, 2H); 4.49 (d, J = 6.00 Hz, 1H); 4.61 (d, J = 5.96 Hz, 1H); 6.10 (s,1H); 6.19 (dt, J₁ = 12.44 Hz, J₂ = 3.60 Hz, 1H); 6.50-6.60 (br m, 3H);6.81-6.86 (m, 1H); 7.02-7.23 (m, 3H); 7.25-7.32 (m, 2H); 7.34-7.38 (m,2H); one exchangeable proton. Compound 3 Racemic

  Compound 3 3-(3,5-Difluorophenyl)-2-{4-[(E)-3-(4- fluoromethylpiperidin-1- yl)propenyl]phenyl}-4-methyl-2H- chromen-7-ol (DMSO-d₆, 500MHz); 1.25-1.29 (m, 2H), 1.65 (d, J = 11.3 Hz, 3H), 1.91-1.96 (m, 2H),2.12 (s, 3H), 2.91 (d, J = 11.1 Hz, 2H), 3.1 (d, J = 6.4 Hz, 2H), 4.26(d, J = 5.5 Hz), 1H), 4.35 (d, J = 5.7 Hz, 1H), 6.13 (br s, 1H), 6.17(d, J = 2.3 Hz, 1H), 6.26-6.32 (m, 1H), 6.42 (dd, J₁ = 8.4 Hz, J₂ = 2.3Hz, 1H), 6.48 (d, J = 15.9 Hz, 1H), 7.11 (d, J = 6.7 Hz, 2H), 7.18 (dt,J₁ = 18.7 Hz, J₂ = 9.4 Hz, 1H), 7.24 (d, J = 8.5 Hz, 1H), 7.29 (d, J =8.2 Hz, 2H), 7.37 (d, J = 8.2 Hz, 2H), 9.7 (s, 1H). Compound 4 Racemic

  Compound 4 3-(3,5-Difluorophenyl)-2-[4-[(E)-3-[(3R)-3-(fluoromethyl)pyrrolidin-1- yl]prop-1-enyl]phenyl]-4-methyl-2H-chromen-7-ol (DMSO-d₆, 500 MHz); 1.4-1.49 (m, 1H); 1.82-1.93 (m, 1H);2.12 (s, 3H); 2.32-2.4 (m, 1H); 2.4-2.5 (m, 2H); 2.5 (m, 2H); 3.2 (d, J= 6.29 Hz, 1H); 4.28 (d, J = 6.29 Hz, 1H); 4.38 (d, J = 6.36 Hz, 1H);6.14 (s, 1H); 6.17 (d, J = 2.31 Hz, 1H); 6.3 (dt, J₁ = 15.8 Hz, J₂ =6.42 Hz, 1H); 6.41 (dd, J₁ = 8.41 Hz, J₂ = 2.3 Hz, 1H); 6.5 (d, J = 15.9Hz, 1H); 7.09 (br d, 2H); 7.15-7.21 (m, 1H); 7.24 (d, J = 8.43 Hz, 1H);7.29 (d, J = 8.17 Hz, 2H); 7.37 (d, J = 8.2 Hz, 2H); two protons weremerged with dmso. Compound 5 Racemic

  Compound 5 3-(3,5-Difluorophenyl)-4-methyl-2-[4-[(E)-3-[(3R)-3-methylpyrrolidin-1-yl]prop-1-enyl]phenyl]-2H-chromen-7-ol (DMSO-d₆, 500 MHz); 1.06 (d, J =6.75 Hz, 3H); 1.40-1.50 (m, 1H); 2.02-2.10 (m, 1H); 2.12 (s, 3H);2.24-2.36 (m, 1H); 2.40-2.48 (m, 1H); 2.90-2.98 (m, 2H); 3.11 (dd, J₁ =9.90 Hz, J₂ = 7.60 Hz, 1H); 3.55 (d, J = 6.75 Hz, 2H); 6.13 (s, 1H);6.20 (d, J = 2.35 Hz, 1H); 6.33 (dt, J₁ = 15.85 Hz, J₂ = 6.80 Hz, 1H);6.44 (dd, J₁ = 8.40 Hz, J₂ = 2.40 Hz, 1H); 6.66 (d, J = 15.85 Hz, 1H);7.04-7.18 (m, 3H); 7.25 (d, J = 8.45 Hz, 1H); 7.33 (d, J = 8.20 Hz, 2H);7.40 (d, J = 8.20 Hz, 2H); one exchangeable proton. Compound 6 Racemic

  Compound 6 3-(3,5-Difluorophenyl)-2-{4-[(Z)-3-(3-fluoromethylazetidin-1- yl)propenyl]phenyl}-4-methyl-2H- chromen-7-ol(DMSO-d₆, 500 MHz); 2.13 (s, 3H); 2.7-2.9 (m, 1H); 3.02 (t, J = 6.5 Hz,2H); 3.3-3.4 (m, 4H); 4.47 (d, J = 6.18 Hz, 1H); 4.57 (d, J = 6.17 Hz,1H); 5.58 (dt, J₁ = 12.11 Hz, J₂ = 6.2 Hz, 1H); 6.17 (s, 1H); 6.20 (d, J= 2.39 Hz, 1H); 6.43 (dd, J₁ = 8.44 Hz, J₂ = 2.41 Hz, 2H); 7.1-7.15 (m,2H); 7.16-7.27 (br, 4H); 7.35 (d, J = 8.2 Hz, 2H); one exchangeableproton. Compound 7 Racemic

  Compound 7 3-(3,5-Difluorophenyl)-2-[4-[3-[3-(fluoromethyl)azetidin-1-yl]prop-1-ynyl]phenyl]-4-methyl-2H-chromen-7-ol (DMSO-d₆, 500 MHz); 2.11 (s, 3H);2.73-2.82 (m, 1H); 3.17 (t, J = 6.76 Hz, 2H); 3.41 (t, J = 7.51 Hz, 2H);3.49 (s, 2H); 4.50 (d, J = 6.14 Hz, 1H); 4.60 (d, J = 6.14 Hz, 1H); 6.1-6.2 (m, 2H); 6.43 (d, J = 2.2 Hz, 1H); 7.08-7.14 (br, 2H); 7.14-7.20 (m,1H); 7.24 (d, J = 8.44 Hz, 1H); 7.34 (d, J = 8.1 Hz, 2H); 7.40 (d, J =8.1 Hz, 2H), one exchangeable proton. ^(#)Chirality at 2^(nd) positionof 2H-chromene ring.

The biological activities of the compounds of the present disclosure wasdetermined via the following assays:

Selective Estrogen Receptor Degradation (SERD) Assay:

The SERD activity for the test compounds was evaluated in MCF-7 cellsharboring ER wild type, and MCF-7 cells harboring mutant ER (WT/D538Gand WT/Y537S). Briefly, cells were plated in phenol red free RPMI1640medium supplemented with 5% charcoal stripped fetal bovine serum. Theseeding density was 40000 cells/well for MCF-7 WT and MCF-7 D538G and50000 cells/well MCF-7 Y537S in a 48-well plate. Following overnightincubation, the cells were treated with varying concentrations of testcompounds (final concentration: 1000 nM to 0.01 nM, 0.1% DMSO) for 4days. The cells were lysed using PBS supplemented with 1 mM EDTA, 0.5%Triton X-100, 5 mM NaF, 6 M urea, and 1× Protease inhibitor cocktail.The lysate was analyzed for ER alpha protein using western blot. For thewestern blot, the cell lysate (12.5-40 μg total protein) was resolved ona 10% SDS PAGE and transferred onto a PVDF membrane. The blots wereblocked using 5% skim milk powder in 0.1% PBS-T for 1 hour at roomtemperature, followed by co-incubation with rabbit anti-human β-actinantibody and rabbit anti-human ERα antibody for 2 hours at roomtemperature. The blots were subsequently probed with anti-rabbit IgG-HRPconjugate as secondary antibody for 1 hour at room temperature. Theblots were developed by using West Pico Super Signal Chemiluminescencesubstrate and the bands were processed for densitometry analysis usingImage Lab software (BioRad version 6.0.0). ERα band intensity wasnormalized to that housekeeping protein for respective samples. The % ERremaining was calculated by normalizing against the vehicle control (as100%).

MCF-7 Cell Growth Inhibition Assay:

The anti-proliferative activity of the test compounds were evaluated ina growth inhibition assay. Briefly, MCF-7 cells harboring ER wild type(Wt), and MCF-7 cells harboring mutant ER (WT/D538G and WT/Y537S) wereseeded at a density of 1000 cells/well in 96-well plate in phenol redfree RPMI1640 medium supplemented with 10% charcoal stripped fetalbovine serum. The cells were incubated overnight at 37° C. and 5% CO2followed by the addition of varying concentrations of the test compoundsin DMSO. Final concentration of DMSO in the well was 0.5%. Following aseven-day incubation of cells with the test compounds, cell viabilitywas assessed using a Prestoblue™ reagent. The percent inhibition of cellproliferation was calculated by normalizing the cell viability using avehicle control as 0% inhibition of proliferation.

The result of the ER-α degradation assay of the compound of Formula Iand its closely related compounds at 1 nM concentration was as shownbelow in Table 1.

TABLE 1 ER-α degradation assay % of ER-α Compound remaining at 1 nMFormula I 41 Compound 2 100 Compound 3 100 Compound 4 100 Compound 5 85Compound 6 80.8 Compound 7 100

As apparent from the table above, the compound of Formula I showedsignificantly more ER-α degradation in MCF-7 cell line (Wt type) ascompared to structurally close compounds including its regioisomerCompound 2. It was surprisingly found that changing the position ofhydroxyl group in the compound of Formula I from the 7^(th) position tothe 6^(th) position on the chromene moiety, as in Compound 2, led to thecomplete loss of the SERD activity. Compounds 3, 4, and 5, which haveeither 6- or 5-membered heterocycloalkyl rings in the side chain hadnegligible SERD activity (more than 80% of ER remaining), whereas thecompound of Formula I which has a 4-membered azetidine ring in the sidechain showed good degradation of ER (only about 41% of ER remaining). Itwas also surprisingly found that changing the double bond in the sidechain of the compound of Formula I to a triple bond as in Compound 7 ledto complete loss of SERD activity. Moreover, Compound 6, which isgeometric isomer (Cis-isomer) of a compound of Formula I, showed verynegligible SERD activity.

Thus, the compound of Formula I has all the optimum structuralattributes required for a potent selective estrogen receptor degrader.The inventors of the present invention further resolved the compound ofFormula I into its S and R stereoisomers. The S isomer is presented asFormula Ia and the R isomer is presented as Formula Ib in the currentdisclosure.

The compound of Formula Ia and compound of Formula Ib were furthertested for their anti-proliferative activity in growth inhibition assay(MCF-7 cell line) and for ER-α degradation. The results are provided asshown in Table 2 below:

TABLE 2 Comparison of in-vitro potency of compound of Formula Ia withits R enantiomer, compound of Formula Ib ER-α degradation assay Wt MCF-7Growth % of Inhibition ER-α Y537S D538G assay (IC₅₀; nM) remaining IC₅₀IC₅₀ IC₅₀ Compound Wt Y537S D538G at 1 nM (nM) (nM) (nM) Formula Ia 0.36.0 2.2 16.3 0.3 19.6 12.7 Formula Ib 69.9 ~1000 345.4 81.5 10.2(>1000)     (>1000)    

As evidenced from the table above, the compound of Formula Ia showedmultifold superior activity over the compound of Formula Ib in MCF-7growth inhibition assay (wild type), wherein the compound of Formula Iais about 233 times more potent than compound of Formula Ib. The compoundof Formula Ia also showed similar trends in a mutated cell line (Y537S &D538G). The compound of Formula Ia was also found to be multifold morepotent in the ER-α degradation assay as compared to its R-enantiomer,wherein the compound of Formula Ia is about 34 times more potent thancompound of Formula Ib in ER-α degradation (wild type). The R enantiomerhas shown no ER-α degradation in mutated cell line (Y537S & D538G).

Pharmacokinetic Study:

In drug discovery it is very important to have good oral bioavailabilityand pharmacokinetic (PK) profile of the compound in order to become agood drug candidate for oral administration. Many compounds havingpotent in vitro efficacy fail to show therapeutic efficacy in humanbody. The lack of therapeutic efficacy of these compounds is mainly dueto their inappropriate pharmacokinetic properties, such as lowbioavailability, short half-life, rapid metabolism, or rapid clearanceresulting in a short duration of action. Therefore a good drug candidateshould have not only good potency, but also a good pharmacokineticprofile. The pharmacokinetic profile of the compounds of Formula Ia andIb were determined as per the procedure provided below.

PK profiles were evaluated in female SD rats (n=2) following a singleoral dose administration. Rats were fasted for 12-13 hours prior todosing and feed was provided 2 hours after oral treatments. Doseformulations of compounds were prepared in Tween 80 (0.4% v/v of thetotal volume of suspension) in 0.5% carboxymethyl cellulose (CMC).Treatments were given at 50 mg/kg, p.o. To evaluate the plasmaconcentration, 0.2 mL blood was drawn from the rats through theretro-orbital plexus at 0.25, 1, 4, 8, 24, 48 and 72 hour time pointspost-treatment in Eppendorf tubes containing sodium heparin as ananticoagulant (100 IU/mL), and centrifuged immediately at 8500 rpm for 7min at 4° C. Plasma was separated and stored at −70° C., until analysis.The analysis of samples was performed as per the method given below:

Working Calibration Standard and Quality Control Standards Preparation:

Working calibration standards were prepared having 0.10, 0.20, 2.00,4.00, 8.00, 12.00, 16.00, and 20.00 μg/mL concentrations. Qualitycontrol standards were prepared at concentrations of 0.30, 3.00, 10.00,15.00 and 60.00 μg/mL.

Preparation of Working Internal Standards (WIS):

Accurately weighed 5 mg of Carbamazepine was transferred into a 100 mLvolumetric flask, dissolved and diluted up to the mark with diluent(0.1% formic acid in water: acetonitrile 30:70 v/v). A WIS of 5 μg/mLwas prepared from the above solution.

Sample Processing:

5 μL of working standards and quality controls were spiked into 95 μL ofblank plasma. 25 μl from the above spiked standards and quality controlswere aliquoted into microcentrifuge tubes. 25 μL of study samples werealso aliquoted into a micro centrifuge tube. 5 μL of WIS was added inzero standards to linearity and study samples except blank. The sampleswere further vortexed for 10-15 seconds. 150 μL of Milli Q water wasadded to all prepared samples and vortexed for 10-15 seconds. Sampleswere loaded on a preconditioned cartridge (preconditioning was done with1 mL methanol followed by 1 mL Milli Q water). Cartridges were washedwith 2×1 mL of Milli Q water. Elution was done with a 1 mL elutionsolution {acetonitrile: 0.1% formic acid solution (70:30)} in HPLCvials.

LC-MS/MS Method:

Chromatographic separation was achieved on Chiral pack ID (250*4.6 mm,5μ) with a flow rate of 1.25 mL/min with splitter and injection volumeof 20 μL. The sample cooler was maintained at 10° C. The column oventemperature was set to 30° C. The mobile phase consisted of:

Mobile phase A: 1.47 g of ammonium bicarbonate into 1 L of water, 1 mlDEA, pH 9.2±0.1 Mobile phase B: methanol,

wherein mobile phase A was mixed with mobile phase B in the ratio of30:70 v/v, respectively. The retention time of the compound of FormulaIa, compound of Formula Ib and the internal standard were about 8.7, 7.6and 4.2 min, respectively. The overall chromatographic run time was 16minutes.

Detection was performed by tandem mass spectrometry (TSQ Quantum,Discovery MAX, Thermo Electron Corporation) and peak areas wereintegrated using LCquan software version 2.9 QF1. The detector was seton MRM mode where transition of 477.900 m/z→272.982 m/z (CE 20) wasmonitored for compound of Formula Ia or for compound of Formula Ib, andtransition of 237.058 m/z→194.003 m/z (CE 16) was monitored for theinternal standard.

The results of the pharmacokinetic study were as shown below in Table 3.

TABLE 3 PK Studies of compound of Formula Ia and Formula Ib PK in rat at50 mg/kg p.o. dose T_(max) C_(max) AUC_(last) AUC_(inf)_obs T_(1/2)Compound (h) (ng/mL) (hr*ng/mL) (hr*ng/mL) (h) Formula Ia 4.00 343 75829804 26 Formula Ib 4.00 119 1795 2356 10

As apparent from above table, compound of Formula Ia showed a superiorpharmacokinetic profile when compared to its R-enantiomer. The C_(max)of the compound of Formula Ia was found to be about 2.9 fold greaterthan that of the compound of Formula Ib. Similarly the AUC_(last) of thecompound of Formula Ia was found to be about 4.2 fold greater than thatof the compound of Formula Ib.

In-Vivo Efficacy of Compound of Formula I and Compound of Formula Ia:

The efficacy of the compound of Formula I and the compound of Formula Iawere evaluated in female athymic nude mice harboring subcutaneousMCF7-Y537S xenografts. Dose formulations of the compound of Formula Iand the compound of Formula Ia were prepared in Tween 80 (0.4% v/v ofthe total volume of suspension) in 0.5% w/v carboxymethylcellulose (CMC)and administered once daily orally for 28 days. Two perpendiculardiameters of tumor were measured with digital Vernier caliper. Tumorvolume (V) was calculated using the following equation: V=(a²×b)/2,where “a” is the width of the tumor (small diameter), and “b” the length(large diameter), both in millimeters. Tumors were monitored twiceweekly and compared with a vehicle treated group. The results of thestudy were as shown below in Table 4. Both the compounds, of Formula Iaand of Formula I, showed significant reduction in the tumor growth whencompared to the vehicle group (see FIG. 1). The results showed thatcompound of Formula Ia at 50 mg/kg and the racemic compound of Formula Iat 100 mg/kg dose levels showed similar tumor growth inhibition (TGI) ascompared to the vehicle group with 56% and 57% TGI, respectively.

TABLE 4 In-vivo efficacy of compound of Formula Ia and compound ofFormula I in MCF7-Y537S mice xenografts % Tumor growth inhibitioncompared to vehicle group # Dose after 28 days Formula Ia 50 mg/kg,p.o., 28 days 56 Formula I 50 mg/kg, p.o., 28 days 46 Formula I 100mg/kg, p.o., 28 days 57

In summary, the compounds of the present disclosure showed betterpotency in in-vitro assays, for example in an. ER-α degradation assay,than their closely related compounds. The compounds of presentdisclosure also showed good in-vivo efficacy in an MCF7-Y537S xenograft.The compounds of the present disclosure, particularly the compound ofFormula Ia, showed a good pharmacokinetic profile and thus can besuitable for oral administration. The compounds of the presentdisclosure or their pharmaceutically acceptable salts can be formulatedin oral dosage forms and can be used for the treatment of diseases whichare related to modulation of ERs, such as ER-positive breast cancer.

What is claimed is:
 1. A compound3-(3,5-difluorophenyl)-2-[4-[(E)-3-[3-(fluoromethyl)azetidin-1-yl]prop-1-enyl]phenyl]-4-methyl-2H-chromen-7-olrepresented by Formula I:

or a pharmaceutically acceptable salt thereof.
 2. A compound(2S)-3-(3,5-difluorophenyl)-2-[4-[(E)-3-[3-(fluoromethyl)azetidin-1-yl]prop-1-enyl]phenyl]-4-methyl-2H-chromen-7-olrepresented by Formula Ia:

or a pharmaceutically acceptable salt thereof.
 3. A pharmaceuticalcomposition comprising a compound of claim 1 and a pharmaceuticallyacceptable carrier, diluent, or excipient.
 4. A pharmaceuticalcomposition comprising a compound of claim 2 and a pharmaceuticallyacceptable carrier, diluent, or excipient.
 5. A method of treatment of acancer in a human being, the method comprising administering aneffective amount of the compound of claim 1 to a human in need thereof,wherein the cancer is selected from the group consisting of breastcancer, endometrial cancer, brain cancer and ovary cancer.
 6. The methodof treatment of claim 5, wherein the cancer is breast cancer.
 7. Amethod of treatment of a cancer in a human being, the method comprisingadministering an effective amount of the compound of claim 2 to a humanin need thereof, wherein the cancer is selected from the groupconsisting of breast cancer, endometrial cancer, brain cancer and ovarycancer.
 8. The method of treatment of claim 7, wherein the cancer isbreast cancer.
 9. The compound of claim 2, wherein the content ofcompound of Formula Ib

is less than 25%, less than 20%, less than 15%, less than 10%, less than5%, less than 1%, less than 0.5%, less than 0.1, less than 0.05%, lessthan 0.01% w/w or absent with respect to the compound of Formula Ia. 10.The compound of claim 2, wherein the enantiomeric ratio of the compoundof Formula Ia is greater than 75:25, greater than 80:20, greater than85:15, greater than 90:10, greater than 95:5, greater than 99:1 or is100:0.
 11. The pharmaceutical composition of claim 3, wherein an amountof S-enantiomer of Formula I present in the composition is at least 75%of a total amount of enantiomers of Formula I present in thecomposition.
 12. The pharmaceutical composition of claim 11, wherein theamount of S-enantiomer of Formula I present in the composition is atleast 99% of the total amount of enantiomers of Formula I present in thecomposition.
 13. The pharmaceutical composition of claim 12, wherein theamount of S-enantiomer of Formula I present in the composition is 100%of the total amount of enantiomers of Formula I present in thecomposition.